Fertilisation

Question 1

overview of fertilisation

Answer 1

.

Question 2

Describe the Acrosome

Answer 2

Developed from Golgi apparatus.
Bounded by membranes.
Contins various enzymes.
Membrane bound initially.

Changes occur to acrosomal membrane which allow contents to be released.

Question 3

When/how does go through Sperm maturation

Answer 3

Sperm undergo maturation in the epididymis (under stimulation by androgens)

Further fluid absorption (100-fold).
Secretions include fructose, proteins and glycoproteins.
Results in transfer of proteins into and onto the sperm plasma membrane; increase in membrane fluidity.

Question 4

What do sperm acquire during maturation?

Answer 4

They acquire ability to swim progressively
More rigid flagellum
cAMP content of tail increases

Question 5

When are sperm able to fertilise an egg?

Answer 5

Once undergone capacitation.

Sperm are not able to fertilise an egg until they have undergone capacitation

Question 6

WHat is capacitation induced by?
When does capacitation occur?

Answer 6

Induced by various components in female tract

This takes place predominantly within the female tract (proteolytic enzymes, cholesterol ‘sinks’ and higher ionic strength)

Question 7

What are two main characteristics do fully capacitated sperm exhibit?

Answer 7

1) hyperactivated motility
2) changes in sperm plasma membrane properties that subsequently allow the acrosome reaction to happen. (i.e further membrane changes)

Question 8

Why capaitation of sperm?

Answer 8

Why?
Enables sperm more responsive to signals from oocyte.
Sperm able to undergo the acrosome reaction which is essential for fertilisation

Question 9

WHat follows on from capacitation?
WHat is capacitation of sperm?

Answer 9

Acrosome reaction follows on from capacitation.

What?
Stripping or modification of surface glycoproteins
Changes in surface charge; reduction in membrane stability through loss of cholesterol and formation of lipid rafts

Question 10

How does capacitation occur?

Answer 10

How?
Mechanisms not fully understood; active research
Sperm cytoplasm becomes more alkaline - increase in bicarbonate conc.
Higher pH increases calcium permeability and hence intracellular calcium concentration. Subsequenct second messenger activation.
Results in increased adenylate cyclase activity and therefore cAMP production.
Activation of spermatozoal protein kinase A (PKA).
Downstream phosphorylation (including flagellum proteins); signalling pathways.

Question 11

Hyperactivated sperm motility

Answer 11

Once sperm undergone compacitiation they become hyperactivated.

Involves high amplitude, asymmetrical beating of the flagellum

Helps the sperm meet its target

Aids sperm penetration of the zona pellucida

Question 12

What are the difference movements of hyperactivted sperm?

Answer 12

The circling high-curvature (C) and helical (H) motility patterns correspond to the transitional trajectories and retain a sufficient symmetry to permit forward sperm movement. The star spin (S) type of motility, characterized by very low linearity, was generally recognized and appeared as an important predictor of success in in-vitro fertilization. Human sperm hyperactivation is not constant and, even over the course of 1 s, spermatozoa change their motility pattern (biphasic, B).

Helps sperm meet egg.

Question 13

Finding the egg

Answer 13

In humans, over 99% spermatozoa do not even enter the cervix

Survival times: oocytes 6-24 hours after ovulation; sperm 24-48 hours in female tract

Sperm swim through uterus and into oviduct; cilia may help

Oocyte and cumulus cells release chemoattractants to aid sperm

Once fertilised Oocyte moves down the oviduct by muscular contractions and beating cilia

Question 14

Acrosome reaction
WHen does it happen?

Answer 14

Described as the ‘terminal phase of the capacitation process’

Only occurs when sperm meets the egg (therefore doesnt happen to all sperm).

Occurs after sperm head binds to zona pellucida; zona proteins responsible for inducing reaction. Induced by proteins on zona pellucida.

Sperm acrosome swells; the acrosome membrane fuses with the sperm plasma membrane. Results in acrosomal contents being released.

Acrosomal vesicle undergoes exocytosis
Release of hyaluronidase and exposure of acrosin on sperm membrane.

Question 15

Fertilisation- generalised acrosomal process

Answer 15

1. Sperm makrd contact with egg
2. Acrosome reacts with zona pellucica (releases contents).
3. Acrosome reacts with perivitelline space
4. Plasma membranes of sperm and egg fuse.
5. Sperm nucleus enters eg
6. Cortical granules fuse with egg plasma membrane, which renders the vitelline layer impenetrable to sperm. Sperm nucleus enters egg.

Question 16

Zona pellucida - describe the 4 glycoproteins

Answer 16

The zona pellucida is composed primarily of four glycoproteins: ZP1, ZP2, ZP3 and ZP4

ZP1 is a structural protein; cross-links others
ZP2 contains sperm-binding domain necessary for oocyte-sperm recognition and penetration of the zona pellucida. Also responsible for the primary block to polyspermy
ZP3, complexed with ZP4, is involved in primary sperm-egg binding.

Question 17

6 steps of Gamete binding

Answer 17

Hyaluronidase from acrosome digests cumulus cells and exposes acrosin, a protease on inner membrane of sperm

Primary binding: Sperm membrane binds zona pellucida via ZP3 (complexed to ZP4) and a species-specific complementary binding partner on the sperm, (species specific).

Secondary binding: Sperm inner acrosomal membrane binds to zona pellucida via ZP2 on zona and acrosin on sperm

Acrosin digests zona pellucida and oocyte plasma membrane

Adhesion of sperm equatorial region and oocyte membrane

Penetration of sperm head into oocyte

Question 18

Oocyte activation and sperm/oocyte fusion.

Answer 18

Following sperm/oocyte fusion, PLCζ is released into the cytoplasm of the oocyte, where it facilitates the hydrolysis of membrane-bound PIP2to DAG and IP3, triggering Ca2+release from intracellular Ca2+stores, leading to Ca2+induced Ca2+release and oocyte activation.

Question 19

Oocyte activation - calcium concentration.

Answer 19

Increase in cytosolic calcium concentration; calcium oscillations
Stimulates cortical granule release (includes Ovastacin - essential for block of polyspermy)
Protein kinase C (PKC) stimulates phosphorylation of other proteins essential for development of the conceptus

Question 20

Block to polyspermy - how is polyspermy prevented?

Answer 20

Ca2+ stimulates fusion of zygotic cortical granules with oocyte plasma membrane and release of contents into perivitelline space

Enzymes including Ovastacin act on zona pellucida which hardens; inactivation of sperm receptors through ZP2 cleavage: Block to polyspermy

Question 21

Completion of fertilisation: Calcium

Answer 21

Calcium pulses stimulate resumption of meiosis; second polar body is extruded and female pronucleus forms

Question 22

Completion of fertilisation: Sperm

Answer 22

Sperm nucleus decondenses; protamines are replaced by histones; male pronucleus forms

Question 23

Completion of fertilisation: Pronuclei

Answer 23

Pronuclei come together; DNA replication occurs; pronuclear membranes break down and replicated chromosomes align on mitotic spindle ready for first cleavage division

Question 24

Syngamy

Answer 24

Combination of two genomes.

Question 25

Pre-implantation development

Answer 25

Initial cleavage occurs symmetrically and getting smaller
16 cell stage = morula
After divisions not symmetrical.

Inner cell mass become embryo.

Question 26

Zygote transport and implantation

Question 27

What are the main characteristics of capacitated sperm?

Question 28

What is the acrosome reaction and what stimulates it to take place?

Question 29

How is polyspermy blocked?